KillerRed，一种基因编码的荧光蛋白光敏剂介导的光动力疗法对白血病细胞的影响。
The effects of photodynamic therapy on leukemia cells mediated by KillerRed, a genetically encoded fluorescent protein photosensitizer.
机构信息
Department of Pathogenic Microbiology & Immunology, School of Basic Medical Sciences, Xi'an Jiaotong University Health Science Center, 76 West Yanta Road, Xi'an, 710061, People's Republic of China.
出版信息
BMC Cancer. 2019 Oct 7;19(1):934. doi: 10.1186/s12885-019-6124-0.
BACKGROUND
Leukemia is a cancer of blood and bone marrow cells, causing about 300,000 deaths worldwide. Photodynamic therapy (PDT) is a promising alternative for the treatment of malignant tumors. KillerRed is a genetically encoded red fluorescent protein photosensitizer (PS). In this study, we aimed to investigate the effects of KillerRed-mediated PDT on chronic myelogenous leukemia K562 cells, acute monocytic leukemia NB4 cells, and acute monocytic leukemia THP1 cells.
METHODS
KillerRed was expressed in Escherichia coli cells, purified by Q-Sepharose column, and confirmed by western-blotting. The PDT effect on cell proliferation was evaluated by Cell Counting Kit-8 (CCK-8). Cell apoptosis was determined by PE Annexin V/7-AAD staining and flow cytometry. The distribution of KillerRed in leukemia cells was detected by confocal laser scanning microscopy (CLSM) and western-blotting. The ROS generation was measured by flow cytometry.
RESULTS
Pure KillerRed was obtained with a yield of about 37 mg per liter of bacterial cells. KillerRed photodynamic inactivated the leukemia cells in a concentration-dependent manner, but exhibited no obvious dark toxicity. PDT mediated by KillerRed could also induce apoptotic response (mainly early apoptosis) in the three cell lines. The CLSM imaging indicated that KillerRed was distributed within the cytoplasm and nuclei of leukemia cells, causing damages to the cytoplasm and leaving the nuclear envelope intact during light irradiation. KillerRed distributed both in the cytosol and nuclei was confirmed by western blotting, and ROS significantly increased in PDT treated cells compared to the cells treated with KillerRed alone.
CONCLUSIONS
Our studies demonstrated that KillerRed-mediated PDT could effectively inactivate K562, NB4, and THP1 leukemia cells and trigger cell apoptosis, and it has potential to be used individually or complementally, in the treatment of leukemia.
背景
白血病是一种血液和骨髓细胞的癌症，导致全球约 30 万人死亡。光动力疗法（PDT）是治疗恶性肿瘤的一种很有前途的替代方法。 KillerRed 是一种遗传编码的红色荧光蛋白光敏剂（PS）。在这项研究中，我们旨在研究 KillerRed 介导的 PDT 对慢性髓性白血病 K562 细胞、急性单核细胞白血病 NB4 细胞和急性单核细胞白血病 THP1 细胞的影响。
方法
KillerRed 在大肠杆菌细胞中表达，通过 Q-Sepharose 柱纯化，并通过 Western-blotting 确认。通过 Cell Counting Kit-8（CCK-8）评估 PDT 对细胞增殖的影响。通过 PE Annexin V/7-AAD 染色和流式细胞术测定细胞凋亡。通过共聚焦激光扫描显微镜（CLSM）和 Western-blotting 检测 KillerRed 在白血病细胞中的分布。通过流式细胞术测量 ROS 的产生。
结果
获得了纯度约为 37mg/L 细菌细胞的纯 KillerRed。KillerRed 光动力以浓度依赖的方式使白血病细胞失活，但没有明显的暗毒性。KillerRed 介导的 PDT 也可以诱导三种细胞系中的凋亡反应（主要是早期凋亡）。CLSM 成像表明，KillerRed 分布在白血病细胞质和核内，在光照下对细胞质造成损伤，而核膜保持完整。Western-blotting 证实 KillerRed 分布在细胞质和核内，与单独用 KillerRed 处理的细胞相比，PDT 处理的细胞中 ROS 显著增加。
结论
我们的研究表明，KillerRed 介导的 PDT 可以有效灭活 K562、NB4 和 THP1 白血病细胞并触发细胞凋亡，它具有单独或互补使用的潜力，可用于白血病的治疗。
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